SUMMARYPurpose: A mutation in the b 1 subunit of the voltagegated sodium (Na V ) channel, b 1 (C121W), causes genetic epilepsy with febrile seizures plus (GEFS+), a pediatric syndrome in which febrile seizures are the predominant phenotype. Previous studies of molecular mechanisms underlying neuronal hyperexcitability caused by this mutation were conducted at room temperature. The prevalence of seizures during febrile states in patients with GEFS+, however, suggests that the phenotypic consequence of b 1 (C121W) may be exacerbated by elevated temperature. We investigated the putative mechanism underlying seizure generation by the b 1 (C121W) mutation with elevated temperature. Methods: Whole-cell voltage clamp experiments were performed at 22 and 34°C using Chinese Hamster Ovary (CHO) cells expressing the a subunit of neuronal Na V channel isoform, Na V 1.2. Voltage-dependent properties were recorded from CHO cells expressing either Na V 1.2 alone, Na V 1.2 plus wild-type (WT) b 1 subunit, or Na V 1.2 plus b 1 (C121W). Key Findings: Our results suggest WT b 1 is protective against increased channel excitability induced by elevated temperature; protection is lost in the absence of WT b 1 or with expression of b 1 (C121W). At 34°C, Na V 1.2 + b 1 (C121W) channel excitability increased compared to NaV1.2 + WT b 1 by the following mechanisms: decreased use-dependent inactivation, increased persistent current and window current, and delayed onset of, and accelerated recovery from, fast inactivation. Significance: Temperature-dependent changes found in our study are consistent with increased neuronal excitability of GEFS+ patients harboring C121W. These results suggest a novel seizure-causing mechanism for b 1 (C121W): increased channel excitability at elevated temperature.

All cells maintain a voltage across their plasma membranes. Only excitable cells, however, can generate action potentials, the rapid, transient changes in membrane potential that spread along the surface of these unique cells. Action potential generation and propagation occurs through, and is regulated by, the function of voltage‐gated ion channels – proteins with ion‐selective pores that span the cell membrane. Ion channels undergo changes in their structural conformation in response to changes in the electrical field across the membrane. These structural changes cause the opening of pores – channels – through which ions can flow down their electrochemical gradient. The charge carried by ions creates an electrical current and rapidly alters the membrane potential with time‐ and voltage‐dependent properties. This rapid, transient membrane potential change is called the action potential. Action potentials transmit information within neurons, trigger contractions within muscle cells, and lead to exocytosis in secretory cells. Key Concepts: All cells maintain a voltage difference across their plasma membranes. Action potentials are all‐or‐nothing, transient changes in membrane potentials of electrically excitable cells that carry important cellular information. Influx of sodium ions through voltage‐gated sodium channels is responsible for the upstroke of the action potential, whereas efflux of potassium ions through voltage‐gated potassium channels is responsible for the falling phase. Propagation of action potentials depends on gating kinetics of ion channels and intracellular and membrane resistances.

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T1-tetramerisation domain of the Kv1.2 potassium channel. We demonstrate that expression of the NaChBac channel returns to near wild-type expression levels. In addition, the channel retains a tetrameric form following purification. We describe the effects of the potassium channel T1 domain on thermal stability and ligand-binding of the NaChBac channel. The recovery of NaChBac expression through expression of an alternative tetramerisation domain is consistent with similar studies on potassium channels and suggests that it is the presence rather than the nature of the tetramerisation domain that is key to channel assembly.